1. Field of the Invention
The present invention relates to novel phenylalanine derivatives that are inhibitors of α4 (including α4β7 and α4β1) mediated adhesion which could be useful in treating conditions such as asthma, diabetes, rheumatoid arthritis, inflammatory bowel disease and other diseases involving leukocyte infiltration to the gastrointestinal tract or other epithelial lined tissues; such as, skin, urinary tract, respiratory airway and joint synovium.
The inhibitors of the present invention could also be useful in treating conditions involving leukocyte infiltration to other tissues including lung, blood vessels, heart and nervous system as well as transplanted organs such as kidney, liver, pancreas, heart and intestine, and blood vessels.
2. Description of the Related Art
The adhesion of leukocyte to endothelial cells or extracellular matrix proteins is a fundamental process for immunity and inflammation and involves multiple adhesive interactions. The earliest events in this process include leukocyte rolling followed by changes in integrin avidity, which lead to subsequent firm adhesion (for reviews see Butcher, Cell 67:1033-1036 (1991); Harlan, Blood 3:513-525 (1985); Hemler, Annu. Rev. Immunol. 8:365-400 (1990); Osborn, Cell 62:3-6 (1990); Shimizu et al., Immunol. Rev. 114:109-143 (1990); Springer, Nature 346:425-434 (1990); and Springer, Cell 76:301-314 (1994)). In response to chemotactic factors, the leukocytes must migrate through two adjacent endothelial cells and into tissues that are composed, in part, of the extracellular matrix protein fibronectin (FN) (see Wayner et al., J. Cell Biol. 105:1873-1884 (1987)) and collagen (CN) (see Bornstein et al., Ann. Rev. Biochem. 49:957-1003 (1980); and Miller, Chemistry of the collagens and their distribution, in “Extracellular Matrix Biochemistry”, K. A. Piez and A. H. Reddi, editors, Elsevier, Amsterdam, 41-78 (1983)). Important recognition molecules that participate in these reactions belong to the integrin gene superfamily (for reviews see Hemler, Annu. Rev. Immunol. 8:365-400 (1990); Hynes, Cell 48:549-554 (1987); Shimizu et al., Immunol. Rev. 114:109-143 (1990); and Springer, Nature 346:425-434 (1990)).
Integrins are heterodimers composed of non-covalently associated subunits, referred to as the alpha (α) and beta (β) subunits (for reviews see Hemler, Annu. Rev. Immunol. 8:365-400 (1990); Hynes, Cell 48:549-554 (1987); Shimizu et al., Immunol. Rev. 114:109-143 (1990); and Springer, Nature 346:425-434 (1990)). To date, 8 integrin β subunits have been identified which can associate with 16 distinct α subunits to form 23 distinct integrins. The α4β1 integrin, also known as VLA-4 (Very Late Antigen-4), is expressed on a variety of cells including lymphocytes, monocytes and eosinophils (see Hemler et al., J. Bio. Chem. 262:11478-11485 (1987); and Bochner et al., J. Exp. Med. 173:1553-1556 (1991)) and may have an important role in the recruitment of these cells during inflammation. VLA-4 is a receptor for vascular cell adhesion molecule-1 (VCAM-1) (Elices et al., Cell 60:577-584 (1990)) and the connecting segment 1 (CS-1), an alternatively spliced region of the FN A chain (Wayne et al., J. Cell Biol. 109:1321-1330 (1989)). The β7 integrin subunit, first cloned by Erle et al. (Erle et al., J. Biol. Chem. 266:11009-11016 (1991)), is expressed only on leukocytes and is known to associate with two distinct α subunits, α4 (Ruegg et al., J. Cell Biol. 117:179-189 (1992)) and αE (Cerf-Bensussan et al., Eur. J. Immunol. 22:273-277 (1992); and Kilshaw et al., Eur. J. Immunol. 21:2591-2597 (1991)).
The α4β7 complex has three known ligands (VCAM-1, CS-1, MAdCAM-1). One ligand which shows unique specificity for α4β7 is Mucosal Addressin Cell Adhesion Molecule-1 (MAdCAM-1) (see Andrew et al., J. Immunol. 153:3847-3861 (1994); Briskin et al., Nature 363:461-464 (1993); and Shyjan et al., J. Immunol. 156:2851-2857 (1996)). MAdCAM-1 is highly expressed on Peyer's patch high endothelial venules, in mesenteric lymph nodes, and on gut lamina propria and mammary gland venules (Berg et al., Immunol. Rev. 105:5-18 (1989)). Integrin α4β7 and MAdCAM-1 have been shown to be important in regulating lymphocyte trafficking to normal intestine (Holzmann et al., Cell 56:37-46 (1989)).
The second ligand for α4β7 is CS-1 (see Guan et al., Cell 60:53-61 (1990); and Wayner et al., J. Cell Biol. 109:1321-1330 (1989)). The cell-binding site within CS-1 is composed of 25 amino acids where the carboxy terminal amino acid residues, EILDVPST, form the recognition motif (see Komoriya et al., J. Biol. Chem. 266:15075-15079 (1991); and Wayner et al., J. Cell Biol. 116:489-497 (1992)).
The third ligand for α4β7 is vascular cell adhesion molecule-1 (VCAM-1), a cytokine inducible protein expressed on endothelial cells (see Elices et al., Cell 60:577-584 (1990); and Ruegg et al., J. Cell Biol. 117:179-189 (1992)). It remains to be unequivocally shown whether MAdCAM-1, VCAM-1 and CS-1 bind to the same site on α4β7. Using a panel of monoclonal antibodies, Andrew et al. showed that α4β7 interaction with its three ligands involves distinct but overlapping epitopes (Andrew et al., J. Immunol. 153:3847-3861 (1994)). VCAM-1 and CS-1 (see Elices et al., Cell 60:577-584 (1990)) are two ligands which are shared by α4β7 and α4β1. In addition, α4β1 is also known to bind to osteopontin, a protein upregulated in arteriosclerotic plaques (see Bayless et al., J. Cell Science 111:1165-1174 (1998)).